Basophils are recruited to inflamed lungs and exacerbate memory Th2 responses in mice and humans.

BACKGROUND: Although the contribution of basophils as inducers or amplifiers of Th2 responses is still debated, prolonged basophil/CD4 T cell interactions were observed in lungs but not lymph nodes (LNs) of parasite-infected mice. However, the impact of basophils on the function of tissue CD4 effector T cells remains unknown. METHODS: Basophils were purified from the lungs of ovalbumin (OVA)-sensitized and OVA-challenged (OVA-immunized) mice or human peripheral blood for in vivo and in vitro functional studies. Pulmonary basophils were adoptively transferred to OVA-sensitized hosts to assess airway inflammation in bronchoalveolar lavage fluid (BALF) and Th2 responses in lung explants and draining LNs. Basophils were co-cultured with effector T cells or Ag-specific naïve T cells alone or in combination with dendritic cells (DCs); IL-4 production was determined by flow cytometry and ELISA. RESULTS: Basophils accumulated in lungs of OVA-immunized mice. Adoptive transfer of basophils to OVA-sensitized hosts enhanced lung IL-4 and IL-13 release while co-administration of OVA further aggravated airway inflammation and Th2 responses in LNs. Mechanistic in vitro studies revealed that pulmonary basophils interacted with lung CD4 effectors, in the absence of DCs, to increase T cell survival and Th2 cytokine expression at the single cell level but amplified OVA-loaded DC-driven Th2 differentiation. Finally, human basophils augmented in vitro IL-4 expression in effector memory CD4 T cells that include CRTH2(+) cells through IL-4 and TCR-independent pathways. CONCLUSIONS: Basophils may worsen Th2 inflammatory disorders through direct interactions with pathogenic CD4 T cells as well as by enhancing DC-induced Th2 cell development.

Hematopoietic progenitor cells are innate Th2 cytokine-producing cells.

Typically, hematopoietic stem/progenitor cells (HSPCs) reside within the bone marrow (BM) where they give rise to all hematopoietic populations. However, HSPCs also constantly egress from the BM into the blood and circulate through the peripheral tissues where upon encounter with inflammatory stimuli and epithelial cell-derived cytokines they rapidly release very high levels of Th2 cytokines/chemokines and differentiate into Th2 effector cells. The novel concept of the dual function of HSPCs as hematopoietic precursors and potent Th2 cytokine producers has important clinical implications in various inflammatory conditions, including allergic diseases.

Adjuvant activity of pollen grains.

BACKGROUND: The induction of an immune response to a biologically inert soluble protein requires an adjuvant. Here we have examined whether intact grains of pollen display such adjuvant effect, accounting for the immunogenic activity of pollen protein allergens that are devoid of intrinsic pro-inflammatory/adjuvant property. METHODS: Human monocyte-derived dendritic cells (DCs) were cultured with intact grains of grass or ragweed pollen for 48 h. The state of DCs maturation was analyzed by FACS and their cytokine production by ELISA. T cell priming activity of DCs was examined in co-cultures with naïve cord blood-derived CD4(+) T cells. RESULTS: Contact with grains of pollen induced a distinct maturation program in immature DCs. Pollen up-regulated the expression of CD54, CD80, CD83, CD86, HLA-DR, CCR7, and CD40 on DCs. Moreover, CCR5 expression was up-regulated by pollen but suppressed by LPS. In sharp contrast to LPS-stimulated DCs, pollen-treated DCs did not produce cytokines [interleukin (IL)-10, IL-12, tumor necrosis factor (TNF)-alpha] but retained the ability to secrete high levels of these cytokines upon simulation with soluble CD40 ligand and interferon (IFN)-gamma. Pollen-primed DCs strongly stimulated the proliferation of allogeneic naïve CD4(+) T cells and promoted their development into effector cells producing high levels of IL-5 and IL-13 together with moderate levels of IFN-gamma and IL-4. CONCLUSION: Intact grains of pollen induce activation and maturation of DCs in vitro. Similar mechanisms may be effective in vivo, suggesting that pollen grain is not only an allergen carrier but also acts as an adjuvant in the induction phase of the allergic immune response.

Role of CD47 in the induction of human naive T cell anergy.

We recently reported that CD47 ligation inhibited IL-2 release by umbilical cord blood mononuclear cells activated in the presence of IL-12, but not IL-4, preventing the induction of IL-12Rbeta(2) expression and the acquisition of Th1, but not the Th2 phenotype. Here we show that in the absence of exogenous cytokine at priming, CD47 ligation of umbilical cord blood mononuclear cells promotes the development of hyporesponsive T cells. Naive cells were treated with CD47 mAb for 3 days, expanded in IL-2 for 9-12 days, and restimulated by CD3 and CD28 coengagement. Effector T cells generated under these conditions were considered to be anergic because they produced a reduced amount of IL-2 at the single-cell level and displayed an impaired capacity 1) to proliferate, 2) to secrete Th1/Th2 cytokines, and 3) to respond to IL-2, IL-4, or IL-12. Moreover, CD47 mAb strongly suppressed IL-2 production and IL-2Ralpha expression in primary cultures and IL-2 response of activated naive T cells. Induction of anergy by CD47 mAb was IL-10 independent, whereas inclusion of IL-2 and IL-4, but not IL-7, at priming fully restored T cell activation. Furthermore, CD28 costimulation prevented induction of anergy. Thus, CD47 may represent a potential target to induce anergy and prevent undesired Th0/Th1 responses such as graft vs host diseases, allograft rejection, or autoimmune diseases.

Bidirectional negative regulation of human T and dendritic cells by CD47 and its cognate receptor signal-regulator protein-alpha: down-regulation of IL-12 responsiveness and inhibition of dendritic cell activation.

Proinflammatory molecules, including IFN-gamma and IL-12, play a crucial role in the elimination of causative agents. To allow healing, potent anti-inflammatory processes are required to down-regulate the inflammatory response. In this study, we first show that CD47/integrin-associated protein, a ubiquitous multispan transmembrane protein highly expressed on T cells, interacts with signal-regulator protein (SIRP)-alpha, an immunoreceptor tyrosine-based inhibition motif-containing molecule selectively expressed on myelomonocytic cells, and next demonstrate that this pair of molecules negatively regulates human T and dendritic cell (DC) function. CD47 ligation by CD47 mAb or L-SIRP-alpha transfectants inhibits IL-12R expression and down-regulates IL-12 responsiveness of activated CD4(+) and CD8(+) adult T cells without affecting their response to IL-2. Human CD47-Fc fusion protein binds SIRP-alpha expressed on immature DC and mature DC. SIRP-alpha engagement by CD47-Fc prevents the phenotypic and functional maturation of immature DC and still inhibits cytokine production by mature DC. Finally, in allogeneic MLR between mDC and naive T cells, CD47-Fc decreases IFN-gamma production after priming and impairs the development of a Th1 response. Therefore, CD47 on T cells and its cognate receptor SIRP-alpha on DC define a novel regulatory pathway that may be involved in the maintenance of homeostasis by preventing the escalation of the inflammatory immune response.

CD47 ligation selectively inhibits the development of human naive T cells into Th1 effectors.

The CD47 Ag, also named integrin-associated protein, was recently reported to regulate the production of IL-12 by human monocytes and dendritic cells. The present study shows that CD47 ligation by CD47 mAb in primary cultures of cord blood mononuclear cells inhibits IL-12-driven Th1 cell development, as revealed by the cytokine secretion profile at restimulation and IFN-gamma production at the single-cell level. F(ab')(2) fragments of CD47 mAb or the synthetic peptide 4N1K, corresponding to the CD47 binding site of thrombospondin, display the same activity. CD47 engagement does not change the phenotype of IL-12-primed cells from Th1 to Th2 or affect IL-4-induced Th2 cell development. Moreover, CD47 mAb inhibits IL-12- but not IL-4-induced IL-2 production as well as IFN-gamma in primary cultures, which was correlated with a decrease of the IL-12Rbeta2 chain expression. Inclusion of exogenous IL-2 at priming corrects IL-12R expression as well as the inhibition of Th1 cell development. The data thus underline the role of IL-2 in Th1 cell development and further suggest that targeting IL-2 and IL-12 simultaneously may have some therapeutic advantage in Th1 autoimmune diseases.

Human monocyte-derived dendritic cells induce naive T cell differentiation into T helper cell type 2 (Th2) or Th1/Th2 effectors. Role of stimulator/responder ratio.

The subset of dendritic cells (DCs) and the nature of the signal inducing DC maturation determine the capacity of DCs to generate polarized immune responses. In this study, we show that the ability of human monocyte-derived DCs (myeloid DC(1)) to promote T helper type 1 (Th1) or Th2 differentiation was also found to be critically dependent on stimulator/responder ratio. At a low ratio (1:300), mature DCs that have been differentiated after inflammatory (Staphylococcus aureus Cowan 1 or lipopolysaccharide) or T cell-dependent (CD40 ligand) stimulation induced naive T cells to become Th2 (interleukin [IL]-4(+), IL-5(+), interferon gamma) effectors. Th2 differentiation was dependent on B7-CD28 costimulation and enhanced by OX40-OX40 ligand interactions. However, high DC/T cell ratio (1:4) favored a mixed Th1/Th2 cell development. Thus, the fact that the same DC lineage stimulates polarized Th1 or Th2 responses may be relevant since it allows the antigen-presenting cells to initiate an appropriate response for the signal received at the peripheral sites. Controlling the number and the rate of DC migration to the T cell areas in lymphoid tissues may be important for the therapeutic use of DCs.

CD47 engagement inhibits cytokine production and maturation of human dendritic cells.

Upon encounter with bacterial products, immature dendritic cells (iDCs) release proinflammatory cytokines and develop into highly stimulatory mature DCs. In the present study, we show that human monocyte-derived DCs functionally express the CD47 Ag, a thrombospondin receptor. Intact or F(ab')2 of CD47 mAb suppress bacteria-induced production of IL-12, TNF-alpha, GM-CSF, and IL-6 by iDCs. 4N1K, a peptide derived from the CD47-binding site of thrombospondin, also inhibits cytokine release. The inhibition of IL-12 and TNF-alpha is IL-10-independent inasmuch as IL-10 production is down-modulated by CD47 mAb and blocking IL-10 mAb fails to restore cytokine levels. CD47 ligation counteracts the phenotypic and functional maturation of iDCs in that it prevents the up-regulation of costimulatory molecules, the loss of endocytic activity, and the acquisition of an increased capacity to stimulate T cell proliferation and IFN-gamma production. Interestingly, regardless of CD47 mAb treatment during DC maturation, mature DC restimulated by soluble CD40 ligand and IFN-gamma, to mimic DC/T interaction, produce less IL-12 and more IL-18 than iDCs. Finally, CD47 ligation on iDCs does not impair their capacity to phagocytose apoptotic cells. We conclude that following exposure to microorganisms, CD47 ligation may limit the intensity and duration of the inflammatory response by preventing inflammatory cytokine production by iDCs and favoring their maintenance in an immature state.

CD47 ligation induces caspase-independent cell death in chronic lymphocytic leukemia.

Thrombospondin forms a 'molecular bridge' between phagocytic and apoptotic cells through interaction with alphavbeta3/CD36. We report here that engagement of CD47, a newly described thrombospondin receptor, by immobilized monoclonal antibody against CD47 or by thrombospondin induced in all B-cell chronic lymphocytic leukemia clones the cytoplasmic features of apoptosis (cell shrinkage, decrease in mitochondrial transmembrane potential and phosphatidylserine externalization) without the nuclear features (chromatin condensation, appearance of single-stranded DNA, DNA fragmentation and cleavage of poly ADP-ribose polymerase). These cytoplasmic events of apoptosis were not prevented by the addition of caspase inhibitor z-VAD-fmk, or by the presence of survival factors (such as interleukin-4 and gamma interferon) or cell activation. Morphological studies confirmed the integrity of the nucleus and showed swelling of the mitochondria. This caspase-independent death pathway may be relevant to the development of alternate therapeutic strategies in chronic lymphocytic leukemia, which remains an incurable disease.

CD47 ligation selectively downregulates human interleukin 12 production.

Interleukin (IL)-12 plays a key role not only in protective innate and adaptive T helper cell type 1 (Th1) responses but also in chronic inflammatory diseases. We report here that engagement of CD47 by either monoclonal antibody, its natural ligand thrombospondin (TSP), or 4N1K (a peptide of the COOH-terminal domain of TSP selectively binding CD47) inhibits IL-12 release by monocytes. The suppression occurred after T cell-dependent or -independent stimulation of monocytes and was selective for IL-12 inasmuch as the production of tumor necrosis factor (TNF)-alpha, IL-1, IL-6, and granulocyte/macrophage colony-stimulating factor was not inhibited. CD47 ligation did not alter transforming growth factor (TGF)-beta and IL-10 production, and the suppressive effect on IL-12 was not due to autocrine secretion of TGF-beta or IL-10. The IL-12 inhibition was not mediated by Fcgamma receptor ligation, did not require extracellular Ca(2+) influx, but was reversed by two phosphoinositide 3-kinase inhibitors (wortmannin and Ly294002). Thus, engagement of CD47 on monocytes by TSP, which transiently accumulates at the inflammatory site, is a novel and unexplored pathway to selectively downregulate IL-12 response. The pathway may be relevant in limiting the duration and intensity of the inflammatory response, and in developing novel therapeutic strategies for Th1-mediated diseases.

OX40-Mediated cosignal enhances the maturation of naive human CD4+ T cells into high IL-4-producing effectors.

Our previous studies have indicated that naive human CD4+ T cells of neonatal or adult origin may be the initial source of IL-4 which is required for their development into Th2 effectors. In addition to minute amounts of IL-4, anti-CD3/B7.1-activated naive cells also release readily detectable levels of IL-13 and IFN-gamma. The production of IL-4 and IL-13 by naive T cells is differentially regulated by TGF-beta and IL-12. Shortly after activation, naive T cells express surface OX40, a TNF-R family member whose ligand (OX40L) is constitutively expressed on a subset of dendritic cells. Engagement of OX40 on activated naive T cells increases their expression of IL-4 and IL-13, suppresses that of IFN-gamma and promotes their development into Th2-like effectors.

Naive human CD4+ T cells are a major source of lymphotoxin alpha.

It is generally accepted that immunologically naive T cells display a very restricted cytokine production profile consisting mainly of IL-2, which is used as an autocrine growth factor. Here we report that activated naive CD4+ T cells, of neonatal or adult origin, express very high levels of soluble lymphotoxin (LT) alpha (LTalpha3), as determined by ELISA, RNase protection assay, and intracytoplasmic staining. Besides LTalpha3 and IL-2, these cells also produce high levels of TNF-alpha together with significant amounts of IFN-gamma and IL-13. Naive cells also express LTbeta mRNA and the membrane form of LTalpha (LTalphabeta). On average, naive CD4+ T cells secrete four times more LTalpha3 than Th1-like cells, twice more than naive CD8+ T cells, and ten times more than B cells. Thus, naive T cells express a large spectrum of cytokines, mainly of the Th1 type, and the very high levels of LTalpha3/TNF-alpha that they release may play an hitherto unsuspected role in the early stage of T cell-dependent immune responses.

Lymphocyte activation gene-3, a MHC class II ligand expressed on activated T cells, stimulates TNF-alpha and IL-12 production by monocytes and dendritic cells.

Lymphocyte activation gene-3 (LAG-3) is an MHC class II ligand structurally and genetically related to CD4. Although its expression is restricted to activated T cells and NK cells, the functions of LAG-3 remain to be elucidated. Here, we report on the expression and function of LAG-3 on proinflammatory bystander T cells that are activated in the absence of TCR engagement. LAG-3 is expressed at high levels on human T cells cocultured with autologous monocytes and IL-2 and synergizes with the low levels of CD40 ligand (CD40L) expressed on these cells to trigger TNF-alpha and IL-12 production by monocytes. Indeed, anti-LAG-3 mAb inhibits both IL-12 and IFN-gamma production in IL-2-stimulated cocultures of T cells and autologous monocytes. Soluble LAG-3Ig fusion protein markedly enhances IL-12 production by monocytes stimulated with infra-optimal concentrations of sCD40L, whereas it directly stimulates monocyte-derived dendritic cells (DC) for the production of TNF-alpha and IL-12, unravelling an enhanced responsiveness to MHC class II engagemenent in DC as compared with activated monocytes. Thus similar to CD40L, LAG-3 may be involved in the proinflammatory activity of cytokine-activated bystander T cells and most importantly it may directly activate DC.

The vitronectin receptor and its associated CD47 molecule mediates proinflammatory cytokine synthesis in human monocytes by interaction with soluble CD23.

The vitronectin receptor, alphavbeta3 integrin, plays an important role in tumor cell invasion, angiogenesis, and phagocytosis of apoptotic cells. CD47, a member of the multispan transmembrane receptor family, physically and functionally associates with vitronectin receptor (VnR). Although vitronectin (Vn) is not a ligand of CD47, anti-CD47 and beta3 mAbs suppress Vn, but not fibronectin (Fn) binding and function. Here, we show that anti-CD47, anti-beta3 mAb and Vn, but not Fn, inhibit sCD23-mediated proinflammatory function (TNF-alpha, IL-12, and IFN-gamma release). Surprisingly, anti-CD47 and beta3 mAbs do not block sCD23 binding to alphav+beta3+ T cell lines, whereas Vn and an alphav mAb (clone AMF7) do inhibit sCD23 binding, suggesting the VnR complex may be a functional receptor for sCD23. sCD23 directly binds alphav+beta3+/CD47(-) cell lines, but coexpression of CD47 increases binding. Moreover, sCD23 binds purified alphav protein and a single human alphav chain CHO transfectant. We conclude that the VnR and its associated CD47 molecule may function as a novel receptor for sCD23 to mediate its proinflammatory activity and, as such, may be involved in the inflammatory process of the immune response.

OX40 costimulation enhances interleukin-4 (IL-4) expression at priming and promotes the differentiation of naive human CD4(+) T cells into high IL-4-producing effectors.

Th2 cell development is critically dependent on the presence of interleukin-4 (IL-4) at priming. The cellular origin and the mechanisms regulating this early production of IL-4 at the site of naive T-cell priming are extensively investigated. We previously reported that anti-CD3-activated and CD28-costimulated naive human CD4(+) T cells themselves release very low but sufficient levels of IL-4 to support their development into high IL-4-producing cells. We show here that ligation of OX40 Ag, a member of the tumor necrosis factor receptor (TNF-R) family, on activated umbilical cord blood CD4(+) T cells upregulates IL-4 production at priming and thereby promotes their development into effector cells producing high levels of the type 2 cytokines IL-4, IL-5, and IL-13. OX40 ligation increases four times the expression of IL-4 mRNA after 48 hours of anti-CD3/B7.1 activation and significantly augments the release of IL-4 and IL-13 in primary cultures. The effects of OX40 costimulation on Th cell differentiation are observed in the presence of optimal and suboptimal CD28 stimulation. Because OX40 ligand is expressed on dendritic cells, the OX40 costimulation pathway may be involved in the physiological regulation of Th cell development by augmenting the differentiation of IL-4-producing cells.

IL-15 promotes IL-12 production by human monocytes via T cell-dependent contact and may contribute to IL-12-mediated IFN-gamma secretion by CD4+ T cells in the absence of TCR ligation.

At inflammatory sites, the number of activated bystander T cells exceeds that of Ag-activated T cells. We investigated whether IL-15, a monocyte-derived cytokine that shares several biologic activities with IL-2, may contribute to bystander T cell activation in the absence of IL-2 and triggering Ag. The addition of IL-15 to cocultures of monocytes and T cells stimulates CD4+ but not CD8+ T cells to produce IFN-gamma. IFN-gamma production requires endogenous IL-12, the production of which in turn is dependent upon CD40/CD154 interactions between CD4+ T cells and monocytes. Indeed, non-TCR-activated CD4+ but not CD8+ T cells express significant levels of CD154. IL-15 may enhance IFN-gamma in this system by up-regulating CD40 expression on monocytes and IL-12Rbeta1 expression on CD4+ T cells. Conversely, using neutralizing anti-IL-15 mAb, we show that the ability of IL-12 to augment IFN-gamma secretion is partly mediated by endogenous IL-15. Finally, in the absence of monocytes, a synergistic effect between exogenous IL-12 and IL-15 is necessary to induce IFN-gamma production by purified CD4+ T cells, while IL-15 alone induces T cell proliferation. It is proposed that this codependence between IL-12 and IL-15 for the activation of inflammatory T cells may be involved in chronic inflammatory disorders that are dominated by a Th1 response. In such a response, a self-perpetuating cycle of inflammation is set forth, because IL-15-stimulated CD4+ T cells may activate monocytes to release IL-12 that synergizes with IL-15 to induce IL-12 response and IFN-gamma production.

Maturation of human neonatal CD4+ and CD8+ T lymphocytes into Th1/Th2 effectors.

The increased susceptibility of neonates to infections has been ascribed to the immaturity of their immune system. More particularly, T cell-dependent responses were shown to be biased towards a Th2 phenotype. Our studies on the in vitro maturation of umbilical cord blood T cells suggest that the Th2 bias of neonatal response cannot be simply ascribed to intrinsic properties of neonatal T cells. Phenotypically, neonatal CD4+ T cells are more immature than their adult CD45RO-/RA+ naive counterparts and they contain a subset (10-20%) of CD45RO-/RA+ CD31- cells which is very low in adults and displays some unique functional features. The activation and maturation of neonatal CD4+ T cells is particularly dependent upon the strength of CD28-mediated cosignal which dictates not only the cytokine profile released upon primary activation but also the response to IL-12. Activation of adult as well as neonatal CD4+ T cells in the context of low CD28 costimulation yields to the production of low levels of only one cytokine, i.e. IL-2. In contrast, strong CD28 costimulation supports the production of high levels of type 1 (IL-2, IFN gamma and TNF beta) and low levels of type 2 (IL-4 and IL-13) cytokines by neonatal T cells. The low levels of naive T cell-derived IL-4 are sufficient to support their development into high IL-4/IL-5 producers by an autocrine pathway. The ability of IL-12 to prime neonatal CD4+ T cells for increased production of IL-4 (in addition to IFN gamma) is observed only when CD28 cosignal is minimal. Under optimal activation conditions (i.e. with anti-CD3/B7.1 or allogenic dendritic cells) the response and the maturation of neonatal and adult naive T cells are similar. Thus the Th2 bias of neonatal immune response cannot be simply ascribed to obvious intrinsic T cell defect but rather to particular conditions of Ag presentation at priming. Unlike CD4+ T cells, neonatal CD8+ T cells strictly require exogenous IL-4 to develop into IL-4/IL-5 producers. Most importantly, anti-CD3/B7-activated neonatal CD8 T cells coexpress CD4 as well as CCR5 and CXCR4 and are susceptible to HIV-1 infection in vitro.

IFN-alpha priming of human monocytes differentially regulates gram-positive and gram-negative bacteria-induced IL-10 release and selectively enhances IL-12p70, CD80, and MHC class I expression.

Administration of IFN-gamma and IFN-alpha may protect or induce autoimmune diseases. Although the in vitro regulation of monokine secretion by IFN-gamma have been extensively studied, the regulatory function of IFN-alpha has not yet been elucidated. We compared IFN-alpha and IFN-gamma, added alone or simultaneously before bacterial stimulation, for the control of monokine release and the expression of costimulatory molecules by human monocytes. Our data show that: 1) IFN-alpha primes monocytes for increased production of IL-10 in response to Staphylococcus aureus Cowan I strain (SAC) but not to LPS, leading to a lack of IFN-alpha priming for TNF-alpha secretion; 2) pretreatment of monocytes with IFN-alpha inhibits LPS- or SAC-induced IL-12p40 production but unexpectedly enhances the release of the biologically active form of IL-12 (IL-12p70); 3) IFN-alpha and IFN-gamma exert an antagonistic effect on LPS- and SAC-induced IL-10 as well as IL-12p40 release, whereas they further enhance IL-12p70 production when added simultaneously; 4) in contrast to IFN-alpha, IFN-gamma primes monocytes to enhance LPS- or SAC-induced TNF-alpha and IL-12 production, but surprisingly, it increases IL-10 production by monocytes following LPS but not SAC stimulation; and finally, 5) IFN-alpha pretreatment selectively up-regulates CD80 and MHC class I expression on monocytes. It is proposed that the outcome of the immune response at the site of inflammation may depend on both the type of bacterial injury (gram-positive or -negative) and of locally produced IFNs, and that the differential and opposite effects of type I and type II IFNs on monocytes may account for the beneficial or detrimental effects of IFN-alpha therapy.

Productive infection of neonatal CD8+ T lymphocytes by HIV-1.

CD8+ T lymphocytes confer significant but ultimately insufficient protection against HIV infection. Here we report that activated neonatal CD8+ T cells can be productively infected in vitro by macrophage-tropic (M-tropic) HIV-1 isolates, which are responsible for disease transmission, whereas they are resistant to T cell-tropic (T-tropic) HIV strains. Physiological activation of CD8-alpha/beta+ CD4- T cell receptor-alpha/beta+ neonatal T cells, including activation by allogeneic dendritic cells, induces the accumulation of CD4 messenger RNA and the expression of CD4 Ag on the cell surface. The large majority of anti-CD3/B7.1-activated cord blood CD8+ T cells coexpress CD4, the primary HIV receptor, as well as CCR5 and CXCR4, the coreceptors used by M- and T-tropic HIV-1 strains, respectively, to enter target cells. These findings are relevant to the rapid progression of neonatal HIV infection. Infection of primary HIV-specific CD8+ T cells may compromise their survival and thus significantly contribute to the failure of the immune system to control the infection. Furthermore, these results indicate a previously unsuspected level of plasticity in the neonatal immune system in the regulation of CD4 expression by costimulation.

Expression and function of OX40 ligand on human dendritic cells.

OX40 ligand (OX40L), a member of the TNF family, was shown to be capable of signaling both the cells on which it is expressed and those expressing OX40, its cognate receptor. Here we show that OX40L is expressed on dendritic cells (DC), the most efficient APC to prime naive T cells. The expression and the functional activity of OX40L were examined by means of mAbs used to stain or cross-link OX40L on 1) freshly isolated human blood DC (bDC) and 2) monocyte-derived DC at different stages of differentiation. These were derived from monocytes cultured either with IL-4 and granulocyte-macrophage CSF (IL-4-Mo-DC) or with IL-4 and granulocyte-macrophage CSF plus TNF-alpha. Both types of Mo-DC expressed OX40L after stimulation through CD40; ligation of OX40L on activated IL-4-Mo-DC enhanced by 4- to 35-fold their cytokine production (TNF-alpha, IL-12 p40, IL-1 beta, and IL-6) and increased CD80, CD86, CD54, and CD40 expression. Stimulation of activated IL-4-Mo-DC through OX40L strikingly enhanced their maturation as evidenced by CD83 up-regulation, CD115 (CSF-1R) down-regulation, and typical morphologic changes. OX40L was constitutively expressed on a subset of bDC, and its ligation slightly enhanced CD40L-stimulated IL-12 production. OX40L was down-regulated after overnight culture and spontaneously reexpressed on a subset of mature bDC (CD83high, CD33high, CD11chigh, CD5+). Thus, the expression of OX40L on DC suggests a physiologic role of this molecule during T cell priming by virtue of its ability to costimulate both T cell and DC activation and differentiation.